POS0255 A PRO-NOCICEPTIVE POPULATION OF NEUTROPHILS INFILTRATE SENSORY GANGLIA AND MEDIATE CHRONIC WIDESPREAD PAIN IN FIBROMYALGIA SYNDROME
Authors: R. Evans, S. Caxaria, P. Celik, A. M. Fuller, S. Bharde, S. Sikandar
Journal: Annals of the Rheumatic Diseases (2023)

<h3>Background</h3> Although the aetiology of chronic widespread pain in fibromyalgia syndrome is unknown and generally recognised as a central pain syndrome, several recent studies point towards a peripheral aetiology. Aberrant activity of immune cells and associated cytokine signalling has also been linked to fibromyalgia syndrome. Neutrophils are polymorphonuclear granulocytes that normally exist in circulation to function as primary mediators of rapid innate host defence, releasing extracellular vesicles as functional mobile units. Surprisingly, neutrophils are found in increased circulatory levels and with enhanced chemotactic and microbicidal properties in fibromyalgia patients. Furthermore, we have recently shown a key role for neutrophils in mediating the spatiotemporal spread of hypersensitivity in a hyperalgesic priming model of chronic widespread pain. Our data also shows a fundamental pro-nociceptive action of neutrophils derived from patients or mice with chronic widespread pain when administered to naïve recipient mice. <h3>Objectives</h3> The mechanisms by which neutrophils can sensitise sensory neurons to cause pain signalling is still unclear but is likely linked to a distinct subpopulation of neutrophils characterised by altered surface protein expression. We aim to identify a specific neutrophil subpopulation that sensitises nociceptive neurons either through cell-cell interactions or mediated via neutrophil-derived extracellular vesicles (NDEVs) to produce chronic widespread pain in fibromyalgia syndrome. <h3>Methods</h3> To identify a pro-nociceptive neutrophil population, we used neutrophils from fibromyalgia patients and pain free controls, characterising proteomic differences between the cohorts by FACS. Furthermore, we have employed functional assays of neutrophil reactivity, including quantification of reaction oxygen species (ROS) production and neutrophil extracellular trap generation (NETosis) to characterise specific functional differences which may responsible their pro-nociceptive capacity. To assess the role of NDEVs in neutrophils pro-nociceptive actions we have isolated and performed phenotypic characterisation of NDEVs from patient and pain-free controls. Finally, we used primary sensory neuron cultures to assess the effect of neutrophils and NDEVs on neuronal excitability using functional assays (calcium imaging and <i>in vitro</i> electrophysiology) to demonstrate the capacity of neutrophils to sensitise peripheral sensory neurons. <h3>Results</h3> Proteomic characterisation of neutrophils derived from patients with fibromyalgia pain vs pain free subjects reveals a distinct population of cells, including altered expression of cell surface markers CD62L and CXCR2. Quantification of ROS generation also reveals changes in cells derived from patients when compared to pain free counterparts, alongside increased NETosis which may reflect specific mechanisms employed by cells to sensitise peripheral neurons following trafficking to DRG. We observe increased NDEV release from patient cells alongside specific phenotypic differences, including an upregulation of surface markers, mirroring what we observe within the parent cells. Moreover, <i>in vitro</i> systems, including electrophysiology recording and calcium imaging reveal increased neuronal responses to both patient cells and patient cell-derived NDEVs. <h3>Conclusion</h3> We show a surprising role of neutrophils, a short-lived immune cell, in the aetiology of chronic widespread pain in fibromyalgia syndrome. Our data demonstrates altered proteomic profiles and distinct phenotypic differences in both circulating neutrophils and NDEVs from patients, supporting an immunological basis for chronic widespread pain in fibromyalgia and a novel mechanism of nociplastic pain. <h3>REFERENCES:</h3> NIL. <h3>Acknowledgements:</h3> NIL. <h3>Disclosure of Interests</h3> None Declared.